Cell Cycle Analysis by Flow Cytometry

For cell-cycle analysis, cells were trypsinized and fixed with 70% ice-cold ethanol for 30 min at 4 °C. After washing twice with PBS (Gibco, Thermo Fisher Scientific Waltham, MA, USA), 5 µL RNAse A (20–40 mg/mL) (Thermo Fisher Scientific, Waltham, MA, USA) was added and incubated for 30 min at 37 °C. After the addition of 200 µL PI (from 50 µg/mL stock solution) (Abcam, Cambridge, UK), the stained cells were analyzed on a BD Accuri C6 Plus Flow Cytometer (BD Biosciences, Franklin Lakes, NJ, USA) using FlowJo software.

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Uebel A., Kewitz-Hempel S., Willscher E., Gebhardt K., Sunderkötter C, & Gerloff D. (2023). Resistance to BRAF Inhibitors: EZH2 and Its Downstream Targets as Potential Therapeutic Options in Melanoma. International Journal of Molecular Sciences, 24(3), 1963.

Publication 2023

PBS RNAse A BD Accuri C6 Plus Flow Cytometer

Cell cycle Cells Cold Ethanol Pi 200 Rnase

Corresponding Organization : Martin Luther University Halle-Wittenberg

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Variable analysis

independent variables

Cell fixation with 70% ice-cold ethanol for 30 min at 4 °C

dependent variables

Cell cycle analysis

control variables

Washing cells twice with PBS
Incubation with RNAse A (20–40 mg/mL) for 30 min at 37 °C
Staining with propidium iodide (PI) from 50 µg/mL stock solution
Analysis on a BD Accuri C6 Plus Flow Cytometer using FlowJo software

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